A Study on Pesticide Residues in Fishery Products : Analytical Method Optimization, Monitoring, and Risk Assessment = 수산물 중 잔류농약 연구 : 분석법 최적화, 모니터링 및 위해성 평가

국문 초록 (Abstract)

본 연구는 어류, 갑각류 및 패류 내 잔류농약 모니터링을 위한 효과적인 농약 동시 다성분 분석법을 개발 및 검증하고, 국내산 수산물 중 잔류농약 모니터링 및 위해성 평가를 수행하였다. Alachlor를 포함한 44종의 농약에 대한 동시 분석법을 GC-µECD 및 GC-MS/MS를 이용하여 total ion chromatograms（TIC）, matrix effect（ME） 및 회수율 시험을 통해 비교하였다. LC-MS/MS 대상 acetamiprid 등 24종 농약의 분석법 최적화를 위하여 회수율 시험을 통해 비교하였다. 잔류농약 분석을 위한 시료 전처리 방법은 QuEChERS（Quick, Easy, Cheap, Effective, Rugged, Safe）법 및 pesticide analytical method（PAM）를 참고 및 변형하여 확립하였다. 추출 용매는 0.1% acetic acid 포함된 acetonitrile을 사용하였으며, 정제 과정에서 GC-MS/MS 대상 성분은 d-SPE（C18 흡착제）법과 florisil cartridge법을 비교하였다. LC-MS/MS 대상 성분은 EN15662 및 AOAC법을 비교하였다. GC-MS/MS 분석법 최적화 단계에서 기존 d-SPE 정제에 비해 florisil cartridge를 활용한 정제 시 복잡한 matrix를 가진 수산물 시료에서 불순물과 색소를 더 효과적으로 제거하고 matrix effect를 감소하는 것으로 확인되었다. LC-MS/MS 대상 성분의 경우 EN15662법이 AOAC법에 비해 높은 회수율을 나타내었다. 분석법 검증은 Codex 가이드라인（CAC/GL 40）을 근거로 하였다. 검출한계는 0.25~3 ng/g, 정량한계는 3~10 ng/g 범위로 나타났다. 검량선은 matrix-matched로 작성하였으며, 결정계수（R²）가 0.98을 초과하는 우수한 선형성을 나타내었다. 정확도와 정밀도는 spike 농도를 3가지（LOQ, 10×LOQ, 50×LOQ）로 달리하여 시험하였으며, 회수율은 60-120%, 상대 표준 편차는 20% 이내로 Codex 가이드라인 기준을 만족하였다. 검증된 분석법은 어류, 갑각류 및 패류를 포함하여 전국 9개 지역에서 채취한 300건의 수산물 시료 분석에 적용되었다. 분석된 시료 중 잉어, 메기, 참돔, 뱀장어, 미꾸라지, 잉어 및 농어에서 8종 농약이 검출되었다. 검출된 농약으로는 boscalid, DDT, isoprothiolane, lufenuron, oxadiazon, pendimethalin, thifluzamide 및 triflulain이다. 검출된 성분의 잔류량은 모두 10 ng/g 이하로 검출되었다. 위해성 평가를 위해 일일 추정 섭취량（EDI）, 일일 섭취 허용량（ADI）값을 사용하여 만성 노출 평가를 수행했다. 또한 급성 기준 참고치（ARfD）가 설정된 농약에 대해서는 급성 노출 평가를 하였다. 수산물 섭취에 따른 평가와 실제 식이 섭취에 따른 농·축·수산물 통합 위해성 평가를 위해 농산물 및 축산물의 섭취량도 함께 사용하였으며, 만성 및 급성 노출 평가에 대한 수준은 5% 미만으로 모두 매우 안전한 것으로 확인되었다. 이 결과는 국내 양식 수산물 섭취에 따른 농약 위해도가 매우 낮은 수준이며, 통합 위해성 평가에서도 낮은 위해도를 나타내어 소비자 건강에 잠재적인 위해를 초래하지 않는 수준인 것으로 확인되었다.

다국어 초록 (Multilingual Abstract)

This study aimed to develop and validate an effective multi-residue analytical method for pesticide residue monitoring in fish, crustaceans, and shellfish, and to conduct monitoring and risk assessment of pesticide residues in domestically produced fishery products. A simultaneous analytical method for 44 pesticides, including alachlor, was optimized using GC-µECD and GC-MS/MS through comparisons of total ion chromatogram （TIC）, matrix effect （ME）, and recovery tests. For the 24 pesticides analyzed by LC-MS/MS, including acetamiprid, recovery tests were performed to optimize the analytical method. Sample preparation procedures for pesticide residue analysis were established by modifying the QuEChERS （Quick, Easy, Cheap, Effective, Rugged, Safe） method and the Pesticide Analytical Manual （PAM）. Acetonitrile containing 0.1% acetic acid was used as the extraction solvent. For GC-MS/MS target compounds, the d-SPE （C18 sorbent） method and Florisil cartridge clean-up method were compared, whereas the EN15662 and AOAC methods were evaluated for LC-MS/MS target compounds. During the optimization of the GC-MS/MS analytical method, Florisil cartridge clean-up was found to remove impurities and pigments more effectively and to reduce matrix effects compared with conventional d-SPE clean-up in fishery product samples with complex matrices. For the LC-MS/MS target compounds, the EN15662 method exhibited higher recoveries than the AOAC method. Method validation was performed according to the Codex guidelines （CAC/GL 40）. The limit of detection （LOD） ranged from 0.25-3 ng/g, and the limit of quantification （LOQ） ranged from 3-10 ng/g. Matrix-matched calibration curves exhibited excellent linearity, with coefficients of determination （R²） greater than 0.98. Accuracy and precision were evaluated at three fortification levels （LOQ, 10×LOQ, and 50×LOQ）. Recoveries ranged from 60% to 120%, and relative standard deviations （RSD） were within 20%, satisfying the Codex guideline criteria. The validated analytical method was applied to the analysis of 300 seafood samples collected from nine regions across the country, including fish, crustaceans, and shellfish. Among the analysed samples, eight types of pesticides were detected in carp, catfish, red seabream, eel, loach, crucian carp and sea bass. The pesticides detected were boscalid, DDT, isoprothiolane, lufenuron, oxadiazon, pendimethalin, thifluzamide and triflulain. For risk assessment, chronic dietary exposure and risk assessments were conducted using the estimated daily intake （EDI） and acceptable daily intake （ADI）. In addition, acute exposure assessments were performed for pesticides with established acute reference dose （ARfD） values. To compare exposure based on fishery product consumption alone with actual dietary exposure, consumption data for agricultural and livestock products were also incorporated into an integrated risk assessment of agricultural, livestock, and fishery products. The results of both chronic and acute exposure assessments were below 5%, indicating a very low level of risk. These findings suggest that pesticide exposure through the consumption of domestically farmed fishery products poses a negligible risk to consumers. Furthermore, the integrated risk assessment also demonstrated a low level of concern for human health, indicating that the detected pesticide residues are unlikely to pose a potential health hazard to consumers.

목차 (Table of Contents)

• Ⅰ. Introduction 1
• Ⅱ. Materials and methods 12
• 1. Optimization of analytical methods for pesticide residues in fishery products 12
• 1) Chemical and materials 12
• 2) Analytical pesticides 13
• 3) Optimization of analyrical methods 18
• (1) GC-MS/MS analysis 18
• (2) LC-MS/MS analysis 23
• (3) Final extraction and purification 26
• 4) Instrumental and analysis conditions 30
• (1) GC-MS/MS and GC-µECD instrumental conditions 30
• (2) LC-MS/MS instrumental conditions 37
• 5) Method validation 41
• 6) Statistical analysis 43
• 2. Monitoring of pesticides residues in fishery products in the Republic of Korea 44
• 1) Sample collection 44
• 2) Sample preparation 46
• 3. Integrated risk assessment of fishery, agricultural, and livestock products 47
• 1) Pesticide residues in fishery, agricultural, and livestock products 47
• 2) Food intake in fishery, agricultural, and livestock products 47
• 3) Chronic exposure 48
• 4) Acute exposure 50
• Ⅲ. Results and discussion 52
• 1. Optimization of analytical methods for pesticide residues in fishery products 52
• 1) Establishment of sample extraction and purification methods 52
• (1) GC-MS/MS 52
• (2) LC-MS/MS 66
• 2) Method validation 69
• (1) Total ion chromatograms 69
• (2) LOD and LOQ 72
• (3) Linearity 79
• (4) Accuracy and precision 86
• 2. Monitoring of pesticides residues in fishery products in the Republic of Korea 97
• 1) Ovrall occurrence of pesticide residues in fishery products 97
• 2) National MRL and potential sources of detected pesticides 102
• 3. Integrated risk assessment of fishery, agricultural, and livestock products 106
• 1) Pesticide residues in fishery, agricultural, and livestock products 109
• 2) Food intake in fishery, agricultural, and livestock products 134
• 3) Chronic exposure 138
• 4) Acute exposure 142
• Ⅳ. Conclusion 146
• Reference 148
• Abstract 163